D W Griffin1, C A Kellogg, K K Peak, E A Shinn. 1. United States Geological Survey, Center for Coastal and Regional Marine Studies, St Petersburg, FL 33701, USA. dgriffin@usgs.gov
Abstract
AIMS: A method for the rapid extraction of fungal DNA from small quantities of tissue in a batch-processing format was investigated. METHODS AND RESULTS: Tissue (< 3.0 mg) was scraped from freshly-grown fungal isolates. The tissue was suspended in buffer AP1 and subjected to seven rounds of freeze/thaw using a crushed dry ice/ethanol bath and a boiling water bath. After a 30 min boiling step, the tissue was quickly ground against the wall of the microfuge tube using a sterile pipette tip. The Qiagen DNeasy Plant Tissue Kit protocol was then used to purify the DNA for PCR/sequencing applications. CONCLUSIONS: The method allowed batch DNA extraction from multiple fungal isolates using a simple yet rapid and reliable assay. SIGNIFICANCE AND IMPACT OF THE STUDY: Use of this assay will allow researchers to obtain DNA from fungi quickly for use in molecular assays that previously required specialized instrumentation, was time-consuming or was not conducive to batch processing.
AIMS: A method for the rapid extraction of fungal DNA from small quantities of tissue in a batch-processing format was investigated. METHODS AND RESULTS: Tissue (< 3.0 mg) was scraped from freshly-grown fungal isolates. The tissue was suspended in buffer AP1 and subjected to seven rounds of freeze/thaw using a crushed dry ice/ethanol bath and a boiling water bath. After a 30 min boiling step, the tissue was quickly ground against the wall of the microfuge tube using a sterile pipette tip. The Qiagen DNeasy Plant Tissue Kit protocol was then used to purify the DNA for PCR/sequencing applications. CONCLUSIONS: The method allowed batch DNA extraction from multiple fungal isolates using a simple yet rapid and reliable assay. SIGNIFICANCE AND IMPACT OF THE STUDY: Use of this assay will allow researchers to obtain DNA from fungi quickly for use in molecular assays that previously required specialized instrumentation, was time-consuming or was not conducive to batch processing.
Authors: Francoise M Blachere; William G Lindsley; James E Slaven; Brett J Green; Stacey E Anderson; Bean T Chen; Don H Beezhold Journal: Influenza Other Respir Viruses Date: 2007-05 Impact factor: 4.380